LILRB1 ligation inhibits immunostimulatory function in allogeneic MLRs that is overcome by CD80 or CTLA-4 blockade or by removal of CD4⁺ CD25⁺ or CD4⁺ CD25⁺ CD127^lo Tregs. (A) Cell populations were irradiated and used to stimulate CFSE-labeled allogeneic T-cell proliferation in MLRs for 7 days. Proliferation was assessed by flow cytometric analysis of CFSE levels in responder CD3⁺ T cells. Results are representative of 4 experiments. (B) Cell populations were irradiated and used to stimulate CFSE-labeled allogeneic T-cell proliferation in MLR in the presence of mAbs specific for CTLA-4 or CD80, or isotype controls for 7 days. Proliferation was assessed as above. Error bars depict the standard deviation of triplicate cultures. Results are representative of 2 experiments. (C) Cell populations were irradiated and used to stimulate CFSE-labeled allogeneic CD4⁺ T cells, from which CD25⁺ cells had been depleted by negative selection, for 7 days. Proliferation was assessed as above. Each panel depicts an individual responder-stimulator allogeneic combination, and error bars indicate SD of triplicate cultures. (D) LILRB1-ligated DCs were irradiated and used to stimulate CFSE-labeled allogeneic CD4⁺ T cells (▬) or allogeneic CD4⁺ T cells from which the CD25⁺ CD127^lo population had been depleted (▭) or were added back to cultures () for 10 days. Each set of 3 values depicts an individual responder-stimulator allogeneic combination, and error bars indicate SD of triplicate cultures.